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目的:通过测定人绒毛膜癌细胞JEG-3中SEPSECS、TRNAU1AP两蛋白的半衰期,分析JEG-3细胞内SEPSECS、TRNAU1-AP两蛋白表达的稳定性。方法:采用蛋白合成抑制剂放线菌酮作用于人绒毛膜癌细胞JEG-3,用MTT法确定放线菌酮对细胞的最适作用浓度,之后用最适浓度的放线菌酮处理JEG-3,利用Western blot检测10个时间点SEPSECS、TRNAU1AP蛋白表达量,确定SEPSECS、TRNAU1AP蛋白半衰期。结果:CHX作用于细胞的最适浓度为0.1μg/mL,与对照组相比,SEPSECS蛋白表达量在4 h、8 h有显著性差异(P<0.01);而TRNAU1AP蛋白表达量在2 h、4 h有显著性差异(P<0.01)。结论:在JEG-3细胞中SEPSECS蛋白的半衰期可能为4 h～8 h,而TRNAU1AP蛋白的半衰期可能为2 h～4 h,前者半衰期明显大于后者,由此,可推测SEPSECS蛋白在JEG-3细胞中表达的稳定性要高于TRNAU1AP蛋白,同时也为进一步分析二者在其他肿瘤细胞的表达情况提供依据。
OBJECTIVE: To determine the stability of SEPSECS and TRNAU1-AP expression in JEG-3 cells by measuring the half-lives of SEPSECS and TRNAU1AP in human choriocarcinoma JEG-3 cells. Methods: Cycloheximide was used as a protein synthesis inhibitor to act on human choriocarcinoma JEG-3 cells. MTT method was used to determine the optimal concentration of cycloheximide to cells. The optimal concentration of cycloheximide was used to treat JEG -3, Western blot was used to detect the expression of SEPSECS and TRNAU1AP at 10 time points to determine the protein half-life of SEPSECS and TRNAU1AP. Results: The optimum concentration of CHX in cells was 0.1μg / mL. Compared with the control group, the expression of SEPSECS protein was significantly different at 4 h and 8 h (P <0.01), while the expression of TRNAU1AP at 2 h , 4 h had significant difference (P <0.01). Conclusion: The half-life of SEPSECS protein in JEG-3 cells may be 4 h to 8 h, while the half-life of TRNAU1AP protein may be 2 h to 4 h, the former half-life is significantly greater than the latter. Therefore, 3 cells than the stability of TRNAU1AP protein expression, but also provide a basis for further analysis of the expression of the two in other tumor cells.